Interleukin-7 (IL-7), a cytokine that plays a key role in the generation, activation and homeostasis of the T-cell compartment, is currently under clinical investigation as a potential therapeutic agent for the immune reconstitution of HIV-1-infected patients. The role of IL-7 in the course of HIV-1 infection is still uncertain. Increased plasma levels of IL-7 have been documented in the course of HIV infection, in association with a reduced expression of the alpha-chain of the IL-7 receptor (CD127) in both CD4+ and CD8+ T cells. The increase in plasma IL-7 was found to correlate with reduced peripheral CD4+ T-cell counts and higher levels of plasma viral load. The most plausible interpretation of these findings is a compensatory effort by the immune system to restore homeostatic levels of CD4+ T cells. However, it is evident that in patients with advanced HIV-1 disease this compensatory effort is insufficient. Thus, it has been suggested that the early administration of IL-7 to patients chronically infected with HIV-1 may help to limit the depletion of CD4+ T cells and/or to promote the regeneration of naive T cells from the precursor pool. Experimental injection of IL-7 in vivo in macaques chronically infected with SIV was shown to increase naive T-cell counts, and to increase proliferation and activation of both CD4+ and CD8+ memory T cells, without augmenting the viral load. A phase I, randomized, placebo-controlled, double-blind trial (IND # BB12069) sponsored by NIAID is currently underway to evaluate the safety of subcutaneous single dose IL-7 in HIV-1-infected subjects who are receiving antiretroviral treatment. [unreadable] Since IL-7 is known to exert anti-apoptotic effects during the course of T-cell development, we have investigated its ability to protect T cells from spontaneous apoptosis in a cohort of HIV-1-infected subjects selected to represent different stages of disease. Recombinant human IL-7 was used over a wide dose range (0.5-50 ng/ml) to determine whether its prosurvival and proliferative activities can be uncoupled. In accordance with previous results, we found that levels of spontaneous apoptosis at baseline were similar in cells from HIV-1-infected patients and HIV-seronegative controls, although they became significantly higher in cells from HIV-1-infected subjects during the course of 6 days of ex vivo culture. Upon addition of exogenous IL-7, a dramatic reduction in the level of spontaneous apoptosis was observed in peripheral blood mononuclear cells (PBMCs) from all the HIV-1-infected patients tested (n = 24), as determined by two independent methods (annexin V binding and caspase 3 activation). The reduction of apoptosis mediated by IL-7 was associated with an increase in the intracellular level of Bcl-2. By contrast, IL-7 had a less consistent, not significant, anti-apoptotic effect on PBMCs from HIV-1-seronegative subjects. To gain further insights into the in vivo significance of IL-7-mediated apoptosis reduction, we analyzed the correlation between the sensitivity to IL-7 and several demographic, clinical and immunological parameters, including age, disease duration, CD4 and CD8 counts and treatment status. A significant inverse correlation was documented between the sensitivity to the anti-apoptotic effect of IL-7 and the circulating CD4 counts, while all the other parameters were not correlated. This correlation was confirmed by comparison of the IL-7 effects in different groups of patients divided according to a defined cut-off value for each demographic clinical and immunological parameter. To verify whether the protective effects of IL-7 occurred on both CD4+ and CD8+ T cells, we purified the two subpopulations from the peripheral blood of HIV-1-infected patients and cultured them ex vivo in the presence or absence of IL-7. These studies demonstrated that IL-7 exerts a protective anti-apoptotic effect of similar magnitude on both T-cell subpopulations from HIV-1-infected subjects. [unreadable] A potential drawback with the in vivo use of immunostimulatory cytokines such as IL-2 and IL-7 is the risk of triggering widespread T-cell activation and proliferation, which could further increase the already high level of immunologic stimulation that occurs in HIV-1 infection and thereby provide more target cells for HIV-1 replication. Thus, we investigated whether the use of IL-7 at concentrations that are sufficient to protect from spontaneous apoptosis was associated with cellular activation and/or proliferation. Intracellular staining with an antibody to the Ki67 antigen, a specific marker of cell cycle progression, showed that IL-7-mediated protection from apoptosis was temporally dissociated from Ki67 expression. Likewise, using the CFSE vital dye or absolute cell counting by flow cytometry, we formally demonstrated that the anti-apoptotic effect of IL-7 occurred in the absence of cell division. Moreover, we measured the levels of endogenous HIV-1 p24 release in cultures of PBMC or purified CD4+ T cells from selected HIV-1-infected patients. We observed that treatment with IL-7, in the absence of mitogenic stimulation, did not result in the induction of significant levels of endogenous HIV-1 replication during short-term ex vivo culture (7-14 days). [unreadable] The anti-apoptotic activity that we documented for IL-7 provides an additional rationale for consideration of this cytokine as an immunotherapeutic agent in the treatment of HIV-1 infection, particularly in patients with more advanced disease who are less likely to regain full immune reconstitution with HAART and whose cells appear to be more sensitive to the anti-apoptotic effects of IL-7.